Current concepts of somatomedin and other biologically related growth factors.

Since the discovery 20 years ago that the growth-promoting effects of somatotropin are mediated through a serum factor(s), research in this area has rapidly expanded. It is the purpose of this review to bring this area of scientific endeavor into perspective. The first part of this review will deal with aspects of total serum somatomedin activity and its biologic actions and measurements in health and disease. The last part of this review will summarize some of the physical and biologic characteristics of the recently purified "somatomedin-like" substances: somatomedin A, B, and C, NSILA-S (nonsuppressible insulin-like activity-soluble in acid ethanol) and MSA (multiplication-stimulating activity).

Circulating alanine production and disposal in healthy subjects.

UNLABELLED: Circulating-alanine production and disposal rates were estimated in eight healthy postabsorptive subjects by means of U-14C alanine and U-14C glucose infusions. The mean circulating-alanine production rate was 368 +/- S.E.M. 28 mumol/min. -1.8(2). Approximately 50 percent of circulating-alanine carbon exchanged rapidly with that of circulating lactate. Approximately 30 per cent of circulating alanine exchanged with protein stores. Other disposal was 29 +/- 2 per cent to circulating glucose and 40 +/- 4 per cent to oxidation. CONCLUSIONS: (1) The carbon moieties of circulating alanine and lactate are freely exchangeable. (2) Assessment of the contribution of alanine to gluconeogenesis will depend on establishing the extent to which the precursor pyruvate carbon is derived from glycolysis or from proteolysis. (3) If the principal pyruvate precursor is glycolysis, then the principal specific function of the glucose-alanine cycle appears to be ammonia transport.

Circulating alanine disposal in diabetes mellitus.

Alanine was selected for study as a representative circulating glucose precursor in relation to the question of the source of the excess circulating glucose in diabetes mellitus. U-14C alanine and U-14C glucose infusions were given to healthy subjects and to subjects with untreated mild maturity, severe maturity, and juvenile diabetes. Comparative studies after a 24-hour fast were made in healthy and in mildly diabetic subjects. The alanine production rate was unaltered by fasting or diabetes. The glucose production rate was unaltered by fasting but increased in diabetes in relation to the severity of the disease. The fractions of alanine-to-glucose and of glucose-from-alanine were increased by fasting. The effect of diabetes was different. The fraction of alanine-to-glucose was much less in mild maturity diabetes than in health, and it was increased only in juvenile diabetes. In all the diabetic groups the glucose-from-alanine fraction was much less than in health. In every group the change in the alanine oxidation rate was reciprocal to that in the alanine-derived glucogenesis rate. The results are consistent with the possibility that the principal source of the excess circulating glucose in diabetes is glycogen.

Isolation of a somatomedin from plasma of rats bearing growth hormone secreting tumors.

We have purified a protein which has somatomedin-like properties from the serum of Wistar-Furth rats bearing a growth hormone producing pituitary tumor (MStT/W15). Activity was measured by a placental insulin and/or somatomedin C radioreceptor assay (SmC-RRA). The serum was initially filtered through Sephadex G-150 equilibrated with 0.1 M NH4HCO3 and 0.02% NaN3. On the G-150 column, radioreceptor insulin (RRI) and radioreceptor somatomedin C (RRSm-C) activities coincided and appeared predominantly in the 160,000 mol wt range with a minor proportion in the 50,000 mol wt range. The pooled active fractions were boiled for 30 min at pH 5.5. After removing denatured protein by centrifugation, the extract was passed through G-50 Sephadex equilibrated with 1% formic acid and 0.15 M NaCl. Sixty to 90% of the SmC-RRA activity in the effluent appeared in the 9000 mol wt range. This material has an isoelectric focusing range of 8.4--9.6, similar to that described for human somatomedin C. On SDS-urea polyacrylamide gel electrophoresis only one protein band was seen. The isolated peptide (rSm) stimulated sulfate uptake in hypophysectomized rat cartilage. The potency of two preparations was variously assayed from 14.0 to 54.7 units/mg. Rat somatomedin was iodinated and purified by absorption on and elution from placental membranes. Eight to 12% of rat [125I]Sm was specifically bound by human placental membranes. Rat [125I]Sm was displaced by hSmC and rSm and human NSILA-S, partially displaced by procine proinsulin and poorly displaced by rat insulin. In preliminary studies, rat [125I]Sm was displaced from receptors on human placental membranes by sera from pituitary tumor bearing rats greater than normal rat sera greater than hypophysectomized rat sera.

Amniotic fluid reactivity detected by somatomedin C radioreceptor assay: correlation with growth hormone, prolactin and fetal renal maturation.

Amniotic fluid was assayed by a radioreceptor assay utilizing 125I-somatomedin C and placental membranes. Growth hormone and prolactin levels were measured by radioimmunoassay at different gestational ages (8-41 weeks). Somatomedin receptor activity, growth hormone, and prolactin reached high levels during early gestation (8-26 weeks) displaying different patterns of appearance which reflect fetal serum levels of these hormones. After 26 weeks gestation all these hormones decreased in concentration. This decrease showed a strong correlation with fetal renal maturation as measured by amniotic fluid creatinine levels.

Characterization of a protein in mid-term human amniotic fluid which reacts in the somatomedin-C radioreceptor assay.

Amniotic fluid contains materials other than insulin which react in a somatomedin C radioreceptor assay using human placental membranes. The material in mid-gestational amniotic fluid which reacted with the somatomedin C radioreceptor assay eluted slightly after albumin from a Sephadex G-150 column equilibrated with 0.1M NH4HCO3. Neither boiling nor treatment of this fraction with 1% formic acid yielded small molecular weight somatomedin-like peptides. Separation of the somatomedin reactive material (Sm RM) from albumin was achieved by gel filtration through Ultrogel AcA44 in 3.1M NH4HCO3. The active product had an apparent molecular weight of 33,000 to 35,000 Daltons; its isoelectric point determined by focusing was between 4.1 and 5.1. The purified amniotic fluid protein displaced somatomedin C in the somatomedin C radioreceptor assay but did not compete with insulin in the insulin radioreceptor assay. Sm RM produced only a slight stimulation of thymidine uptake in human fibroblasts but was inactive in stimulating sulfate uptake in hypox rat costal cartilage. In human fibroblast cultures Sm RM inhibited the stimulation of thymidine uptake induced by human serum and by purified rat somatomedin. When Sm RM was added to the 125I somatomedin C, some of the radioactivity eluted from gel filtration at pH 8.6 was converted to a molecular weight complex of about 43,000. We conclude that the material which we have isolated from mid-gestational amniotic fluid is a protein which may bind somatomedin and make it unavailable to the somatomedin receptor of human placenta and human fibroblasts.